nd C4 plants differ in their proportions of two carbon isotopes C-12 ...Rather than analyzing plant matter in bulk sediment samples David Nel...Using a spooling-wire micro-combustion device to combust the grains a... Because we analyze carbon isotopes in a material unique to grasses (p...This analysis found unequivocal evidence for C4 grasses in southweste...

nd C4 plants differ in their proportions of two carbon isotopes, C-12 and C-13.) Those studies indicated that C4 grasses were present as early as the Early Micocene, about 18 million years ago.

Rather than analyzing plant matter in bulk sediment samples, David Nelson, a postdoctoral researcher in Hu's lab at the time of the study (now a professor at the University of Maryland), analyzed the carbon isotope ratios of individual grains of grass pollen, a technique he pioneered while working with Hu in the lab of biogeochemistry professor Ann Pearson at Harvard University.

Using a spooling-wire micro-combustion device to combust the grains, and an isotope mass spectrometer to determine the relative ratio of C-12 and C-13 in the sample, Nelson and Illinois graduate student Michael Urban analyzed hundreds of individual grains of grass pollen collected from study sites in Spain and France.

"Because we analyze carbon isotopes in a material unique to grasses (pollen) we were able to detect C4 grasses at lower abundances than previous studies," Nelson said.

This analysis found "unequivocal evidence for C4 grasses in southwestern Europe by the Early Oligocene," the authors wrote. This means these grasses were present 32 to 34 million years ago, well before studies indicate atmospheric carbon dioxide levels made their precipitous decline.

"The evidence refutes the idea that low (atmospheric) CO2 was an important driver and/or precondition for the development of C4 photosynthesis," the authors wrote.

"This study challenges that hypothesis and basically says that something else was responsible for the evolution of C4 plants, probably higher temperature or drier conditions," Hu said. With atmospheric carbon dioxide levels now on the increase, he said, "there are also implications about how C3 and C4 plants will fare in the future."

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